In humans, four important glycoprotein hormone heterodimers (LH, FSH, TSH and CG) have identical .alpha. subunits and differing .beta. subunits. Three of these hormones are present in virtually all other vertebrate species as well; CG has so far been found only in primates and in horses.
PCT application WO90/09800, published 7 Sep., 1990, and incorporated herein by reference, describes a number of modified forms of these hormones. One important modification is C-terminal extension of the .beta. subunit of FSH, LH, and TSH by the carboxy terminal peptide (CTP) of human chorionic gonadotropin .beta.-subunit or a variant thereof. Other muteins of these hormones are also described. The CTP-extended .beta. subunit of FSH is also described in two papers by applicants herein: LaPolt, P. S. et al.; Endocrinology (1992) 131:2514-2520 and Fares, F. A. et al.; Proc Natl Acad Sci USA (1992) 89:4304-4308.
The crystal structure of the heterodimeric form of human chorionic gonadotropin has now been published in more or less contemporaneous articles; one by Lapthorn, A. J. et al. Nature (1994) 369:455-461 and the other by Wu, H. et al. Structure (1994) 2:545-558. The results of these articles are summarized by Patel, D. J. Nature (1994) 369:438-439.
PCT application WO91/16922 published 14 Nov., 1991 describes a multiplicity of chimeric and otherwise modified forms of the heterodimeric glycoprotein hormones. In general, the disclosure is focused on chimeras of .alpha. subunits or .beta. subunits involving portions of various .alpha. or .beta. chains respectively. One construct simply listed in this application, and not otherwise described, fuses substantially all of the .beta. chain of human chorionic gonadotropin to the .alpha. subunit preprotein, i.e., including the secretory signal sequence for this subunit.
PCT application WO96/05224 describes single-chain forms of the glycoprotein hormone wherein the .alpha. and .beta. subunits are covalently linked through their N- or C-termini so as to form agonists or antagonists. The linkage can be either .beta.-.alpha. or .alpha.-.beta.. Also described are single-chain forms wherein two .beta. subunits are covalently linked through their termini. Preferred embodiments of such single-chain forms are fusion proteins wherein the linkage is effected through peptide bonds, optionally through additional amino acid sequence forming a "linker."
It has now been found by applicants that single-chain proteins which consist essentially of two .alpha. subunits covalently linked through their N- or C-termini, optionally through a linker moiety can behave as agonists or antagonists of the glycoprotein hormones by interacting with the relevant receptors.